The conformational free energy surface of α-<i>N</i>-acetylneuraminic acid (Neu5Ac, sialic acid) in the space of ring-puckering coordinates was calculated using the metadynamics method. Free energy surfaces in vacuum and with an explicit solvent were calculated in GLYCAM 06 force field. In vacuum three structures are almost equivalently populated, namely, the <sup>2</sup>C<sub>5</sub> chair and the B<sub>3,6</sub>/<sup>2</sup>S<sub>6</sub> and <sup>O</sup>S<sub>3</sub> boat/skew-boat conformations. The B<sub>3,6</sub>/<sup>2</sup>S<sub>6</sub> structure is stabilized by an ionic hydrogen bond between the amide N−H bond and the carboxylic group. However, this structure is unfavorable in a water environment in which the experimentally observed <sup>2</sup>C<sub>5</sub> chair conformation is predicted to be more stable than the other structures. These results indicate that environment significantly influences conformation of Neu5Ac and that Neu5Ac-processing enzymes might modify a conformation of their substrates solely by a changing polarity of the environment. The structure of Neu5Ac bound in influenza neuraminidase (<sup>4</sup>S<sub>2</sub>/B<sub>2,5</sub>) belongs to conformations preferred in a water environment. The free energy penalty of this conformational change was calculated (relative to <sup>2</sup>C<sub>5</sub>) as 10.2 ± 2.0 and 17.3 ± 2.0 kJ/mol for <sup>4,O</sup>B/<sup>O</sup>S<sub>3</sub> and <sup>4</sup>S<sub>2</sub>, respectively. This result indicates that mimicking of the enzyme-bound conformation is likely to be a viable strategy for the design of neuraminidase inhibitors.